One of the most characteristic features of the aging process is the reduced ability of an organism to maintain homeostasis in response to stimuli at either the cellular or organ level. Age- related changes in gene expression, which have been observed in liver and other tissues, could be an important factor in the age- related decrease in cell's ability to respond to hormonal signals and, therefore, to its ability to respond to changes in environmental and metabolic conditions. Because the regulation of transcription is the basic site of control for all cells, this type of cellular change could be important to all cell types. The objective of the research described in this proposal is to determine if the age-related decline in the expression of phosphoenolpyruvate carboxykinase (PEPCK) in liver of rats is due to an alteration in the genetic apparatus of the hepatocyte. The specific aims of the project and the techniques used for achieving them are as follows: 1. To establish that the decline in PEPCK expression is due to a change at the cellular level rather than to an age-related change in the hormonal status of the animal, the expression of PEPCK will be studied in cultures of hepatocytes isolated from adult and old rats. 2. To determine if the ability of the hepatocytes to express PEPCK in response to a hormonal stimulus becomes ineffective as an organism ages, the induction of PEPCK expression by cAMP and/or dexamethasone will be studied in cultured hepatocytes isolated from adult and old rats. 3. To identify the site (cytoplasmic or nuclear) of the cellular deficit in PEPCK expression, I will develop and use a nuclear/cytoplasmic cell-free transcriptional assay prepared from cultured hepatocytes isolated from adult and old rats. 4. To study the role of chromatin structure in the age-related decline in PEPCK expression, I will compare the nuclease hypersensitive sites in the PEPCK gene of the chromatin isolated from cultured hepatocytes obtained from adult and old rats.